It is widely accepted that intracellular calcium release is a major trigger for early development after fertilization. It is unclear how such release is elicited by sperm. Two general hypothesis have been put forth: """"""""the sperm oscillogen"""""""" hypothesis states that a soluble protein from sperm activates calcium release; the alternative is that sperm initiate calcium release and activate eggs through receptor-mediated signal transduction mechanisms comparable to those involved in cellular signaling in other cells. The primary aim of the proposed research is to test rigorously the possible involvement of guanine nucleotide-binding regulatory proteins (G- proteins) in egg activation in the marine worm, Chaetopterus. This will be done by the techniques of immunology, immunocytochemistry and molecular biology. First we will identify, using antibodies and reverse transcription/polymerase chain reaction, the G-proteins in the eggs that could be involved in the regulation of calcium release. Second we will localize these G-proteins by immunocytochemistry to determine whether they can potentially interact with receptors known to mediate some gamete interactions. Third, we will generate reagents to constitutively activate and inhibit the function of G-proteins to determine whether their action is necessary or sufficient for calcium release or other events in egg activation. These studies will have relevance not only for our understanding of the mechanisms by which sperm activate development at fertilization and thus for the potential control of fertility, but also to our understand of cellular signaling in general and hence to our understanding of the cell biology of many disease states.
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